Excess amounts of Ski family of proto-oncoproteins cause oncogenic transformation of normal cells and disruption of the transforming growth factor beta (TGF-beta) signal transduction pathway. Either overexpression of the proto-oncogene c-ski or expression of its oncogenic form v-ski can transform cells. Ski and its related protein SnoN play important roles in regulating the balance between cell proliferation and differentiation. The biological activity of Ski and SnoN depends on its expression levels. Both transcriptional and post-transcriptional mechanisms can modulate the levels of Ski but the biochemical basis for Ski regulation is poorly defined. The principal investigator's objective is to identify the mechanisms by which the steady state levels of Ski and SnoN are regulated post-translationally and how these control mechanisms are disrupted during Ski induced tumorigenesis. The central hypothesis of this proposal is that the activity of the normal Ski protein is controlled post-translationally by both cell cycle and TGF-beta signaling through regulated proteolytic degradation. Loss of cell cycle control and TGF-beta signaling result in stabilization of Ski and hyperproliferation of cells. This hypothesis is based on the principal investigator's preliminary observation that Ski protein is rapidly degraded in the Gl and S phase of the cell cycle and in response to stimulation with TGF-beta. Excessive amounts of Ski can cause an early onset of S phase of the cell cycle and interdict TGF-beta signaling to cell cycle arrest. The hypothesis will be tested by pursuing the following specific aims:: 1) Define the molecular basis for selective degradation of c-Ski during the cell cycle progression; 2) Determine whether Ski degradation is regulated by cell cycle dependent phosphorylation and the mechanism of Ski phosphorylation; 3) How does TGF-beta regulate selective degradation of Ski and SnoN? Is it the same mechanism as cell cycle? Proper levels of Ski and SnoN are crucial for animal development. Results from experiments outlined in this proposal are expected to contribute significantly to our comprehensive understanding of how protein turnover is controlled during cell cycle progression and response to extracellular stimuli. Understanding the biochemical basis for maintaining proper levels of these two proto-oncoproteins will reveal potential targets for therapeutic intervention.